It is known that metal proteinates are absorbed by plants and affect the growth thereof. U.S. Pat. No. 3,873,296 aptly illustrates this phenomenon. However plants need more than just minerals and nutrients such as N-P-K mixes to affect plant growth. It is known that plants produce hormones such as cytokinins, auxins, and gibberellin(s) which also affect plant growth. Inorganic mineral salts are not absorbed intact and translocated as such throughout plants. Thus, they do not provide the minerals to the plant cells as rapidly as metal proteinates. Inorganic mineral salts are not synergistic with plant hormones and their use with plant hormones range from little to no value when compared to the present invention. Before mineral salts can be of value, the plant must alter its composition whereas metal proteinates are absorbed intact and translocated throughout the plant as metal proteinates.
Metal proteinates are actually chelates formed from a soluble bivalent metal salt and two or more protein hydrolysate ligands. By bivalent metal salt is meant a metal having an oxidation state of at least two or higher. For example transition metals may assume more than one oxidation state. In addition many metal salts carry from two to eight, and sometimes more, waters of hydration which are referred to as coordination numbers of the metal. The coordination number and valence may be different; however, the waters of hydration are attached to the metal by coordination bonding with the oxygen atom of the water supplying the necessary electrons. For example a ferrous sulfate salt may exist as such: ##STR1##
When a chelate is formed the anion is replaced and the metal ion is neutralized so that the corresponding chelate has no net electrical charge. In this manner the metal is transported through the plant more easily.
In order for the plants to have the metal ion in the right place at the right time it is important that the stability constant of the metal chelate be just right. Ligands such as citrates and ascorbates are weak complexes and perhaps even chelates but are easily broken down into their component parts. On the other hand, EDTA (ethylenediaminetetraacetic acid) and its salts form such strong chelates that they pass through an organism largely unaffected. The hydrolysis products of proteins seem to form just the right chelates and are referred to as metal proteinates. A chelate or proteinate is a heterocyclic structure which involves, in this case, at least two ligands. A ligand is the protein hydrolysate used to combine with the metal to form the proteinate and consists of a polypeptide, peptide, or naturally occurring amino acid.
Utilizing glycine, the simplist amino acid, a metal proteinate may thus be formed as follows: ##STR2##
However the reaction is not quite that simple. It is important that all excess protons be removed from the protein hydrolysate prior to its reaction with the metal salt. In other words the protein hydrolysate should be on the basic side of its zwitterion state and thus possesses an excess of electrons where the protons have been removed. Glycine then might be represented as: EQU NH.sub.2 CH.sub.2 COOH.revreaction.NH.sub.2.sup.+ CH.sub.2 COO.sup.-
in its zwitterion state. The glycine, or other ligand is preferably brought to a slightly alkaline pH to remove the carboxy proton and render the amino group electronegative and thus reacts with the metal stepwise as follows: EQU (1) 2NH.sub.2 CH.sub.2 COOH+2NaOH.fwdarw.2NH.sub.2 CH.sub.2 COO.sup.- Na.sup.+ +2H.sub.2 O ##STR3## and then upon the addition of more base (NaOH) to a pH between about 7.5 and 10 the amino group is rendered more electronegative and the reaction is completed as follows: ##STR4##
It will be seen that the ligands have replaced the waters of hydration and the base has reacted to remove the protons and form an alkali metal salt. This is for exemplification only and myriad reaction steps could be shown using various bases and ligands as well as different metals. It will be seen from the above why it is important that each proteinate consists of a metal and at least two ligands in order to completely neutralize any electrical charge.
The exact ligand used is not important, amino acids such as lycine, glycine, valine, methionine and the like may be used along with peptides and polypeptides. While hydrolysates with a peptide bond are preferred as ligands, any example utilizing a protein hydrolysate ranging from naturally occurring amino acids to polypeptides is deemed to exemplify the invention in its best mode.
The same is true with metals. While iron, zinc, and copper may constitute the more important metals calcium, manganese, cobalt, molybdenum, and magnesium, may be equally as exemplary.
Plant hormones (phytohormones) are biologically active materials of plant origin that are effective in minute concentrations at sites remote from tissues in which they are formed. Cytokinins, auxins, gibberellin(s), abscisic acid and ethylene are the major classifications of phytohormones.
Kinetin was the first of the active cytokinins (having growth promoting properties) identified and is a 6-furfurylaminopurine having the formula: ##STR5##
Other naturally occurring cytokinins involve:
dimethlallyl amino purine ##STR6## methylamino purine ##STR7## zeatin (methylhydroxymethylallylaminopurine) ##STR8##
Zeatin has been isolated and chemically identified from young kernals of maize, coconut milk, plums, fungus, bacterium, lupin plants and other plants having soluble ribonucleic acid.
One may also find attached to the amino group phenyl, benzyl, n-ethyl, n-propyl, n-butyl and similar groups. ##STR9## Diphenylurea, a synthetic compound, shown above also exhibits cytokinin activity.
Various cytokinins are found in different sources. Dimethylallylaminopurine occurs in soluble ribonucleic acid of many different organisms and is produced by bacterium corynebacterium fasians.
The bacterium and mutations from dimethylallylaminopurine invade green plants such as algae, chlorella, kelp and by secreting the compound produces cytokinin effects.
The dihydro-derivative of zeatin has been isolated from lupin plants and cytokinins have been isolated from the sporophyte of mosses.
The richest natural sources for kinins that have been isolated are seaweed, fruits, and endosperm tissues.
Diphenylura in the presence of casein hydrolysate is distinctively active in cytokinin effects.
Metals and plant hormones are inseparably connected. Dimethylallylaminopurine has been identified with the transfer of ribonucleic acid which combines with serine and tyrosine before these amino acids are incorporated into protein. This explains the cytokinin effect on ribonucleic acid, protein and chlorophyll levels, and indirectly plant growth. Zinc, manganese and iron are all involved in the process of plant growth. Magnesium is of course essential to chlorophyll formation.
Manganese activates the enzyme indoleacetic acid oxidase which controls the distribution of the growth regulators produced from auxins. This enzyme limits the amount of auxin in any area and prevents excessive amounts. It also deactivates auxin in nongrowing areas.
Zinc builds up the auxin hormone just as manganese regulates and controls the supply.
Iron activates an enzyme transport system that controls directions and movement of plant regulators.
Other minerals mentioned such as copper, boron, molybdenum, and magnesium also have important functions in plants.
Auxins greatly magnify the cytokinin effect. Added cytokinins increase mitosis in roots and encourage mitosis in cultural flower anthers.
Cytokinins are strong promoters of bud growth and leaf growth stimulation. Some other effects of cytokinins in plants result in ending dormancy, promote polarity of growth, promote flowering, increase effectiveness of light in said germination, and promote stem elongation.
The growth of a plant is regulated in an orderly way through photosynthesis and respiration. This is accomplished by sun, water, micronutrients such as molybdenum, manganese, zinc, iron and boron and enzymes and by growth hormones such as the naturally occurring auxin. Chemically speaking this auxin is indoleacetic acid. To be produced a mineral such as zinc is required to stimulate the plant enzyme system to refine the amino acid tryptophane into auxin.
Another mineral, manganese, is required to activate the enzyme indoleacetic acid oxidase which controls the distribution of the growth regulation produced by auxin or indoleacetic acid. As mentioned, manganese also aids in regulation of auxin in non-growing regions of the plant.
Auxins, cytokinins, and gibberellin(s) interact in plant growth development and cell division but oppose each other in lateral bud outgrowths.
There are many synthetic chemicals that behave like the naturally occurring auxins produced by plant enzyme systems. In addition to indoleacetic acids, indol-3-butyric acid; naphthaleneacetamide; 2 methyl-1-naphthaleneacetic acid and 2-methyl-1-naphthylacetamide have hormonal activity and may be substituted for the naturally occurring auxins. The synthetic auxins cannot function without zinc, manganese, and other minerals in the same requirement pattern as found with naturally occurring auxins. For best results, the minerals must be in the form of proteinates. As stated earlier the proteinate preferably has a peptide (--CONH--) bond.
One of the important aspects of plant growth and nutrition is nitrogen fixation. Nitrogen can enter biological systems only when it has been combined with other elements such as hydrogen and oxygen. Industrially nitrogen is converted into such compounds as ammonia, nitrate salts, urea or ammonium sulfate. Nature provides a way for nitrogen fixation using the molecular nitrogen gas (N.sub.2) from the air and enzymatically combining it with hydrogen from carbohydrates or natural gas to form ammonia utilizing a nitrogenase. Certain bacteria also act to form ammonia. No substances between nitrogen and ammonia have been isolated, so all the intermediate states must be bound to the nitrogenase.
In the soil fixed nitrogen is employed in the synthesis of biological molecules. A critical structural element is the peptide bond (--CONH--) which links one amino acid to the next; the bond connects a nitrogen atom in one amino acid to a carbon atom in another. Several amino acids may be linked together to form a peptide or polypeptide which will ultimately form a protein.
A metal proteinate not only provides the plant with an essential trace metal but also has a nitrogen fixation sparing effect thus avoiding several steps in nitrogen fixation and allows the plant to absorb ligands containing the peptide bonds directly. This may be accomplished by means of soil application or foliar spray.
Phytohormones may be prepared synthetically or naturally. Cytokinins are primarily available as seaweed extracts. These extracts are diluted with water and used as foliar sprays or applied to the soil.
Kinetin may be prepared synthetically and has essentially the same activity as cytokinin. Gibberellin(s) have also been obtained from seaweed extracts but store less well than cytokinins or kinetin. Auxins have also been prepared from seaweed extracts.
Several beneficial aspects have been attributed to phytohormones including increased crop yields, improved seed germination, increased resistance of plants to frost, fungal and insect attack, increased uptake of inorganic constituents from the soil, reduction in storage losses of fruit and stabilization of chlorophyll. See Blunden, Marine Natural Products Chemistry, Plenum Publishing Corporation, N.Y., N.Y., 1001, pp 337-344.
Phytohormones are known carriers of certain inorganic substances into a plant but the amount of minerals is only a minute fraction of the total mineral requirement for the plant.
According to Brain et al, The Effects of Aqueous Seaweed Extract on Sugar Beet, Proceedings of the Eighth International Seaweed Symposium, University of North Wales, 1974, seaweed extracts are characterized by their high cytokinetic activity. The most important effects of cytokinins are on cell division, cell enlargement, the delaying of senescense and the related transport of nutrients.
One important factor is that cytokinins are very restricted in their movement within the plant, if indeed they move at all from the original site of application. Treated foliar areas act as metabolic sinks and amino acids, phosphates and other substances accumulate in the plant tissues directly under on close to the site of application. For optimal results the cytokinin or other phytohormones should apread throughout the plant. More is involved with phytohormones than the mere mobilization of nutrients, since the delay of senescence of excised plant parts has been demonstrated many times.
The observation that cytokinin treatment augmented the ratio of RNA to DNA, suggested that a critical effect of cytokinins in senescence might be the maintainence of the protein synthesizing machinery, perhaps by regulating RNA synthesis.
Insofar as sugar beets are concerned the translocation or spreading of cytokinin will increase the leaf size, protein content, chlorophyll and leaf life. Hence, the photosynthetic power of the plant would be increased with cytokinin translocation which would result in increased carbohydrate synthesis and increase the stored carbohydrate content of the root.
Aqueous seaweed extracts have successfully been used as fertilizer additives on bananas, gladiolas, tomatoes, peppers, potatoes, corn and oranges with varying degrees of success. Of special interest was the increased uptake of manganese in banana plants. Also of interest was the improved storage of peaches.
Logically, it is mecessary to know the activity of the phytohormone being used so concentration can be regulated and optimal reproducable results obtained.
The class of phytohormones referred to as auxins may be natural or synthetic such as indoleacetic acid or 2.4-dichlorophenoxy acetic acid (b 2.4D.). These hormones are transported within the root from its base to its apex. Natural occurring auxins are not as stable in ambient air as synthetic auxins. Auxins in general move more rapidly to the root tip when applied to cotyledon or leaves. The movement presumably is accompanied with the transport of carbohydrates via the phloem. Since auxins, as contrasted to cytokinins, move more rapidly through the plant they are adapted to the treatment of seeds prior to planting. The consistent application of phytohormones helps reduce the usage of N.P.K. fertilizers by as much as 25%. Optimally, cytokinins, auxins, and gibberellin(s) are applied at a rate of 0.001 to 4.0 grams per acre. Preferrably these phytohormones are utilized as dilute solutions containing on the order of 10-200 ppm of active ingredient and, if used in an alkaline media, are stabilized by a preservative such as sodium benzoate.
The root of a plant contains portions of the best known phytohormones and serve as a center for synthesis. The xylem and phloem being the major circulatory portions of a plant also serve as hormone carriers for those hormones that can be translocated. It has been documented that there are manifold effects of root hormones, expecially cytokinins, on shoot development. These include control of protein and CO.sub.2 metabolism in leaves, enzyme formation in leaves, leaf aging and senescence, elongation of the shoot, stem elongation, lateral shoot development and release of floral bud dormancy, and fruit set.
Environmental influences which affect the root system such as water stress, flooding, excessive heat or cold act not only on water and ion uptake and transport of organic substrates but also on the hormonal flow from root to shoot and vice versa.
From the above discussion of phytohormones it may be seen that there is a complex interaction between nitrogen fixation and uptake, mineral uptake, phytohormone activity and their role in the translocation of ions and nutrients.
In recognizing the importance of a plant as a whole it becomes apparent that manipulation of the entire plant, including shoots, leaves, and roots by phytohormones and metal proteinates with selective field testing offers an almost limitless avenue in plant growth regulation and improvement.